I propose to determine, by direct methods, the regulatory mechanism(s) by which hepatoma tissue culture (HTC) cells control their level of HMG CoA reductase (HMGR) in response to low density lipoproteins, cholesteryl succinate, and 25-hydroxycholesterol. To accomplish this broad goal, I plan to a) isolate and purify antibodies to normal Buffalo rat liver HMGR, b) determine with anti-HMGR whether the increased catalytic activity of HTC cells maintained in medium which contained lipoprotein poor serum is, in fact, the result of an enhanced rate of HMGR synthesis (Ks) or decreased rate constant for degradation (Kd) or both, and c) assess by kinetic analysis the effect of an acute addition of exogenous steroids to HTC cells on the catalytic activity of HMGR and its turnover. In addition, selected characteristics of purified HMGR from cells exposed to 25-hydroxycholesterol will be compared with the enzyme from unsuppressed cultures.